The major current goal of project is to study the biochemistry and cell biology of Macrophage Stimulating Protein (MSP) and its receptor protein tyrosine kinase (RON). Investigations to date suggest that this ligand/receptor pair may affect a number of cells that participate in the response to tissue injury, including skin cells, macrophages and capillary endothelium. Our major findings this year are: 1. Using time-lapse video, we found that MSP stimulates macrophage membrane ruffling and pinocytotic activity. Wortmannin, an inhibitor of phosphoinositol-3kinase, prevents MSP action, suggesting that PI-3K is in the signal transduction pathway. If wortmannin is added 15 min after MSP, membrane motility is not inhibited. Thus, the MSP PI-3K path is a trigger of motility events that do not require continuing stimulation via PI-3K. 2. The receptor for MSP belongs to the large family of growth factor receptor protein tyrosine kinases, which are thought to be activated by ligand-induced dimerization. We found that the primary receptor binding site of the MSP alpha-beta-chain disulfide- linked heterodimer is in the beta-chain. Free beta-chain binds, but does not activate the receptor. This suggests that a secondary binding site, present in the whole molecule, is required for receptor dimerization. 3. Pro-MSP is the single chain precursor of MSP. Proteolytic cleavage of pro-MSP at a single site yields activie MSP. Peritoneal macrophages, one of the target cells of MSP, has two enzymatic activities that cleave pro-MSP. One cleaves at the activation site, the other degrades pro-MSP. Normal serum has an inhibitor of the degrading enzyme. The inhibitor was isolated and identified by N-terminal sequencing as a alpha-1-antichymotrypsin. Control of MSP activity at the locus of macrophage target cells may involve this enzyme inhibitor combination.